Mechanisms responsible for vascular failure of cardiac grafts, especially the development of transplant-associated coronary artery disease (TCAD), remain obscure. In a murine model of heart transplantation, preliminary data show a direct correlation between extent of initial ischemic injury and the degree of neointimal proliferation, more pronounced in allografts yet nevertheless present in isografts. To elucidate alloantigen-independent mechanisms of graft vascular injury caused by preservation, ischemic vessels were shown to exhibit a loss of vasoprotective nitric oxide (NO) and cyclic nucleotides, increased expression of leukocyte adhesion receptors and plasminogen activator inhibitor-1, inhibited expression of plasminogen activator genes, and increased reactive oxygen species. Novel electrochemical sensors show L-arginine or tetrahydrobiopterin deficiency subverts cNOS to make O2-, and that NO nearly vanishes after murine CTX due to rapid quenching by O2-; restoring deficient second messenger pathways (NO,cAMP, cGMP) with substrates, analogs, or type specific phosphodiesterase inhibitors improves acute graft vascular function and causes a striking reduction in TCAD; these benefits were reversed by inhibiting PKA. As adhesion receptor-null grafts (P-selectin, ICAM-1) or those puled with interleukin-1 receptor antagonist (IL-1ra) are protected from early vascular failure following prolonged preservation, the investigators hypothesize that early ischemic injury drives primary vascular failure and accelerates TCAD development, particularly in alloreactive vessels. The aims are (1) to elucidate mechanisms of preservation injury which contribute to primary graft failure or TCAD. Effects of preservation on electrochemical, functional, and histomorphometric endpoints will be studied in control, transgenic, or gene-deficient mice to assess the functional relevance of genes which modulate the cytokine, leukocyte adhesive, fibrinolytic, or reactive oxidant milieu of the graft vasculature; and (2) to determine if attenuating early graft vascular injury can limit primary graft failure and TCAD development, examining the effects of early modulation of NO or cyclic nucleotide pathways, blockade of endothelial-leukocyte interactions, reduction of graft oxidant stress, or inhibition of intravascular thrombosis. An optimal cardiac preservation strategy will then be devised which protects early graft vascular function and reduces the occurrence or severity of TCAD. These experiments will provide new insights into mechanisms by which early ischemic injury leads to early graft failure and TCAD, and proffer new strategies for prevention.